Estradiol Production by Prostaglandin F2alpha

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DESCRIPTION (provided by applicant): The goal of this proposal is to elucidate the molecular and cellular mechanisms that regulate P450 aromatase (P450arom) expression in the rat corpus luteum (CL). The rat CL produces progesterone and estradiol (E2), both of which are essential for reproduction. P450arom, which catalyzes the last step in E2 biosynthesis, is expressed in the rat CL throughout pregnancy until the day before parturition, when it rapidly decreases. The mechanism(s) that control P450arom expression in rat CL and the factors that cause its fall before parturition, are not known. Preliminary studies indicate that prostaglandin F2alpha (PGF2alpha) decreases luteal P450arom mRNA levels and the transcription rate of the cyp19 gene, which encodes P450arom. The proposed study will address the hypothesis that PGF2alpha acts on luteal cells to regulate specific intracellular signaling pathways oriented to decrease cyp19 transcription. We propose to investigate the following specific aims: i) to further characterize the effect of PGF2alpha on cyp19 expression, we intend to determine the specificity of the PGF2alpha effect by using agonists and antagonists of the PGF2alpha receptor and to examine whether PGF2alpha decreases cyp19 promoter activity by using gene reporter constructs, ii) to identify in vivo which transcription factors regulate basal expression of P450arom in the CL and to examine which factors mediate the effect of PGF2alpha on cyp19 transcription. We have found that GATA-4 binds to the P450arom proximal promoter and that such binding is increased by PGF2alpha. Before continuing our experiments to determine whether GATA-4 mediates PGF2alpha-inhibition of P450arom expression, we propose to examine whether GATA-4 regulates P450arom expression in vivo. We also propose to study whether LRH-1, COUP-TF and/or SF-1 are involved in the regulation of luteal P450arom expression as they regulate P450arom expression in granulosa and endometrial cells. Using RT-PCR, Gel Shift, Chromatin Immunoprecipitation, and Western blot, we will examine in vivo and in vitro the expression and binding activity of these factors in luteal cells. Considering that E2 is involved in the development of human pathologies such as breast cancer and endometriosis, the outcome of this project may contribute information that can be the foundation of future studies on tumor cells, especially if we consider that the same cyp19 promoter is used in corpus luteum, endometriotic plaque, and breast tumor.